Carboxymethylcellulose polyethylene glycol compositions for medical uses

ABSTRACT

Compositions comprising carboxypolysaccharides (CPS) including carboxymethyl cellulose (CMC) and polyethylene glycols (PEGs) are provided where the PEG is a PEG-epoxide covalently linked to the CPS. In certain embodiments, the PEG attaches to only one CPS, forming a decorated CPS. In other embodiments, bi-functional PEG molecules are attached to adjacent CPSs, thereby forming a covalently cross-linked composition. Additional embodiments include PEG/CMC compositions where the PEG is a multi-branch PEG and/or a multi-arm PEG. PEG/CMC compositions can be made with desired viscoelastic properties, and such compositions can be used as space-filling materials, load-bearing materials, anti-adhesion compositions, drug delivery vehicles or lubrication of tissues and medical instruments.

CLAIM OF PRIORITY

This Application is a Division of U.S. patent application Ser. No.11/988,048 filed Nov. 28, 2007, entitled “CarboxymethylcellulosePolyethylene Glycol Compositions for Medical Uses,” Samuel Falcone andRichard Berg, inventors, incorporated herein fully by reference.

FIELD OF THE INVENTION

This invention relates to compositions comprising derivatizedcarboxypolysaccharides (CPSs), and in particular, carboxymethylcellulose(CMC). Specifically, this invention relates to CMC derivatized withpolyethylene glycols (PEGs) to form: (1) PEG-decorated CMC, (2) PEGester-linked to CMC under acid catalysis, (3) PEG ester-linked to CMCunder basic catalysis, and (4) PEG-ether linked to CMC or CPSs, and (5)uses thereof as space filling compositions, load-bearing compositions,lubricants, for antiadhesion compositions and for dermal fillers.

BACKGROUND OF THE INVENTION

Carboxymethylcellulose (CMC) is a water soluble, biocompatible andbioresorbable semi-synthesized polysaccharide. The safety ofcommercially available CMC having high purity has been identified andapproved by the Food and Drug Administration (FDA) for incorporationinto many products. CMC is able to react with various polymers by way ofelectrostatic interaction, ionic cross-linking, hydrogen bonding, Vander Waals interactions, and physical interpenetration. Because of itssafety, convenience and diversity of physico-chemical properties, CMChas demonstrated applications in the pharmaceutical, food and cosmeticindustries.

CMC is one type of carboxypolysaccharide (CPS). CPSs have also been usedin the manufacture of implantable polymers. CPSs are polymers made ofsaccharide monomers in which some of the hydroxyl (—OH) groups arereplaced with carboxyl groups (—COOH or COO—). Thus, CPSs such as CMChave some hydroxyl groups and some carboxyl groups present.Carboxylation can permit ionic interaction within a polymer chain or canpermit interaction between polymer chains, thereby forming a gel. Suchgels have been used for a variety of applications, including implantablemedical polymers.

SUMMARY OF THE INVENTION

Aspects of this invention are based on the previously unknownunderstanding that prior implantable polymer materials exhibitedundesirable properties depending upon the tissue in which they wereplaced. For example, in tissues that have a high inherent elasticity,placing a relatively inelastic polymer can result in stresses on thetissue and can lead to tissue damage and deterioration. Thus, werealized that matching elasticities of tissues and of implantablepolymers placed near or within those tissues can improvedbiocompatability of the polymer material.

Other aspects of this invention are based on the novel understandingthat prior compositions containing CMC or other carboxypolysaccharides(CPSs) cannot permit manufacture of implantable polymers having asufficiently controllable or wide range of elasticity and have improvedbiocompatibility.

Thus, we developed new compositions containing CPSs either decoratedwith or cross-linked with bi- or multi-functional poly(ethylene) glycols(PEGs). In general, PEGs useful in the compositions of this inventionwill have a glycidyl ether moiety. The glycidyl ether moiety is anepoxide, which can form covalent bonds with another reactive group viaan addition reaction, without formation of toxic byproducts. Such PEGSare herein termed “PEG-epoxides.” PEG-epoxides may have one epoxidemoiety, two epoxide moieties (“PEG diglycidyl ether; “PEGDGE”) or mayhave three or more epoxide moieties (“multi-branch PEGs” or “multi-armPEGs”). Multi-branch PEGS are PEGS in which in-chain carbon atoms havean epoxide moiety. Multi-arm PEGS are PEGS in which several PEG polymerchains are attached together via a “hub” moiety, with the PEG chainshaving one or more epoxide moieties. It can be appreciated that PEGshaving one or more glycidyl ether moieties can be used to manufacturecompositions in which PEGs can have increased numbers of cross-linkingmoieties.

In one series of embodiments, a bi-functional PEG, PEG diglycidyl ether(PEGDGE), can be reacted with CPSs under basic or acidic catalysis toform compositions having covalent cross-links between CPS polymerchains. It can be appreciated that multi-functional PEGS (e.g.,bi-functional PEGs or PEGDGE; multi-chain PEGs or multi-branch PEGs canbe used to form compositions having CPSs cross-linked with PEGswithin-chain or between-chains by covalent bonds.

In some embodiments, a PEG-epoxide can form a covalent bond with eithera hydroxyl group of the CPS thereby forming an ether linkage. In otherembodiments, a PEG-epoxide can form a covalent bond with a carboxylgroup of the CPS thereby forming an ester linkage.

In some embodiments, basic catalysis of PEG-epoxide and CPS in thepresence of sodium hydroxide (NaOH) can produce compositions havingreduced elasticity compared to prior art CMC compositions. These newmaterials can be characterized by having relatively high ether contentand relatively low ester content. As the relative amount of PEG-epoxideis increased, the elasticity of the derivatized CPS can be decreased ina controllable fashion, permitting manufacture for the first time ofimplantable CPS-containing polymers having a desired and controlledelasticity.

In other embodiments, basic catalysis of a PEG-epoxide and CPS in thepresence of ammonium hydroxide (NH₄OH) can produce materials havingincreased elasticity compared to prior art CPS-containing compositions.In some of these embodiments, the materials are characterized by havingsignificant ester linkages.

In further embodiments, acid catalysis of a PEG-epoxide and CPSs canproduce materials having increased elasticity compared to prior artcompositions of CPSs.

In still other embodiments, if a relatively low amount of a PEG-epoxidecan be used compared to the number of functional groups (hydroxyl groupsand carboxyl groups) on the CPS, the PEG-epoxide can bind to only onefunctional group, thereby “decorating” the CPS monomer withoutsubstantial cross-linking between different CPS molecules. In otherembodiments, using a higher amount of PEG-epoxide can result in eitherintra-chain or inter-chain cross-linking. Increasing the inter-chaincross-linking can increase elasticity of the material. Thus, byadjusting the relative ratios of PEG-epoxide and CPS, and by adjustingthe conditions of catalysis (e.g., NaOH, NH₄OH or acidic catalysis), theelasticity of the resulting polymer material can be finely adjusted tomatch the elasticity of the tissue in which the polymer is to beimplanted.

Additional aspects include use of multi-arm or multi-branch PEGs, whichcan have multiple reactive epoxide moieties thereon. Use of suchmulti-arm and/or multi-branch PEGs can provide gels with morecross-linking per unit of CPS, and therefore higher elasticity thanmaterials made with PEGDGE (a “bi-functional” PEG).

Further, in other aspects, CMC/PEG gels can be made and polymerized insitu without further precipitation and/or purification steps.Embodiments of these aspects can be useful where ease of manufacture isdesirable.

The new compositions find use in a variety of medical and otherapplications. Compositions containing such PEG-CPS linked materials canbe formed into membranes or into gels for antiadhesion uses, eitherduring primary surgery or to decrease re-adhesion during correctivesurgery, for long-term hydrogel implants, and for disk replacement.Manufacture of such membranes, beads, particulates, coatings and gelsand some of their uses has been described in U.S. Pat. Nos. 5,096,997,6,017,301, 6,034,140, 6,133,325, 6,566,345, 6,869,938 and 7,192,984,each patent expressly incorporated herein fully by reference as ifindividually so incorporated.

Compositions of this invention can be useful for delivering drugs totissues. The sites of delivery of drugs using the compositions of thisinvention include, without limitation, skin, wounds, mucosa, internalorgans, endothelium, mesothelium, epithelium. In certain embodiments,buccal, optical, nasal, intestinal, anal, vaginal applications usingcompositions of this invention can be used.

BRIEF DESCRIPTION OF THE FIGURES

The invention will be described with respect to the particularembodiments thereof. Other objects, features, and advantages of theinvention will become apparent with reference to the specification anddrawings in which:

FIG. 1 is a diagram depicting decoration of a CMC polymer with PEGDGEand forming either an ether or an ester bond.

FIG. 2 is a diagram depicting inter-chain cross-linking of CMC polymerswith PEGDGE via two ether linkages of this invention.

FIG. 3 is a graph depicting the ratio G″/G′ (tan δ) vs. frequency forCMC/PEGDGE complexes cross-linked in the presence of NaOH catalysis ofthis invention.

FIG. 4 is a graph depicting the effect of epoxide equivalents on tan δfor CMC/PEGDGE complexes of this invention having different ratios ofCMAG/EP.

FIG. 5 depicts Fourier Transform Infra Red (FTIR) spectrograms ofCMC/PEGDGE complexes of this invention formed under basic conditions inthe presence of NaOH. Curves A, B, C and D represent spectra ofcomplexes having different ratios of CMAG/EP.

FIG. 6 depicts a graph of tan δ vs. frequency of CMC/PEGDGE complexes ofthis invention cross-linked under basic catalysis in the presence ofNH₄OH.

FIG. 7 depicts a graph of FTIR spectrograms for CMC alone (graph A), orCMC/PEGDGE (graph B). The complexes were cross-linked under basiccatalysis in the presence of NH₄OH, showing the characteristic shoulderof ester linkages in graph B (arrow).

FIG. 8 depicts graphs of tan δ vs. frequency of CMC A and CMC/PEGDGEcomplexes of this invention, cross-linked under acidic catalysis andhaving different ratios of CMAG:EP.

FIG. 9 depicts FTIR spectra of acid catalyzed reaction products of CMC Awith PEGDGE. Graph A is of un-reacted CMC A and graph B is of CMCA/PEGDGE reacted in 1% acetic acid with a CMAG/EP ratio of 3.6. Graph Cis of CMC A/PEGDDE reacted in 0.01% citric acid with a CMAG/EP ratio of14.3.

FIG. 10 depicts tan δ vs. frequency of un-reacted CMC B (graph A) andCMC B/PEGDGE complexes of this invention, cross-linked under acidiccatalysis and having a ratio of CMAG/EP of 5.1:1 (graph B).

FIG. 11 depicts FTIR spectra of CMC and CMC/PEGDGE complexes of thisinvention, cross-linked under acidic catalysis and having ratio ofCMAG/EP of 5.1:1.

FIG. 12 depicts a multi-branched PEG-epoxide useful for manufacturingcompositions of this invention.

FIG. 13 depicts a multi-arm PEG-epoxide useful for manufacturingcompositions of this invention.

FIG. 14 depicts a graph of tan δ vs. frequency for two gels, one (graphA) made with CMC C and polyethylene oxide (PEO) and heat-sterilized, andanother (graph B) made with CMC C and PEGDGE and subsequently heatsterilized.

FIGS. 15 a and 15 b depict two embodiments of this invention, in which aPEG/CPS composition is enclosed in a bag.

FIG. 15 a depicts an embodiment of this invention in which a cylindricalbag has a PEG/CPS composition therein.

FIG. 15 b depicts an embodiment of this invention in which a sphericalbag has a PEG/CPS composition therein.

FIGS. 16 a and 16 b depict views of vertebrae with a PEG/CPS compositionin a bag as a replacement for the nucleus pulposus surrounded by anannulus.

FIG. 16 a depicts a top view of a vertebra showing a vertebral body,annulus and a PEG/CPS-filled bag therein as a replacement for thenucleus pulposus.

FIG. 16 b depicts a side view of two adjacent vertebrae, showing anintervertebral space with a PEG/CPS-filled bag as a replacement for thenucleus pulposus surrounded by an annulus.

FIGS. 17 a and 17 b depict views of vertebrae with a plurality ofPEG/CPS-filled bags within an annulus, wherein the plurality of bags isa replacement for the nucleus pulposus.

FIG. 17 a depicts a top view of a vertebra with a plurality ofPEG/CPS-filled bags within an annulus.

FIG. 17 b depicts a side view of two adjacent vertebrae, showing anintervertebral space with a plurality of PEG/CPS-filled bags within anannulus.

FIG. 18 depicts a top view of an embodiment of this invention in whichan intervertebral space has an annulus having a defect and showing aplurality of PEG/CPS-filled bags therein, with one bag occluding thedefect.

DETAILED DESCRIPTION Definitions

The following definitions apply in general to the descriptions thatfollow. In certain cases, however, a term may be defined differently. Inthose cases, the proper definition will be provided.

The term “cross-linking” or “crosslinking” means covalent bonding of twopolysaccharide chains together using a chemical reagent.

The term “decoration” means covalent bonding of a chemical reagent to asingle polysaccharide chain.

The term “CMC” means sodium carboxymethylcellulose. “CMC A” is HerculesCMC grade 7H PH lot #82666. “CMC B” is Hercules grade 9M31F PH lot#90252. “CMC C” is Hercules grade 7H PH lot #92013.

The term “cross-linker” or “crosslinker” means a chemical agent used inthe covalent bonding of two polysaccharide chains to one another.

The term “PEO” means polyethylene oxide, a polymer made up of repeatingunits of compounds containing —(O—CH₂—CH₂)—. PEOs have molecular weightsof greater than about 5000 kDa.

The term “Dalton” or “D” means a unit of molecular mass, where one D isequivalent to the mass of a proton.

The term “PEG” or “polyethylene glycol” means a polymer made ofrepeating units of compounds containing —(O—CH₂—CH₂)— but havingmolecular weights in the range of about 200 Daltons to about 5000 kDa.

The term “epoxide” means an organic functional group with an oxygen atombonded to two adjacent carbon atoms, which has a chemical formula:—CH(O)CH₂. An epoxide is also herein termed a “glycidyl ether.” Anepoxide is a very reactive functional group that can be used incross-linking or decoration reactions.

The term “glycidyl ether” means an organic functional group thatcontains the moiety: —CH(O)CH₂. An example is poly(ethylene) glycoldiglycidyl ether (PEGDGE), a crosslinker used in embodiments of thisinvention. It should be appreciated that multi-branched PEGs andmulti-arm PEGs are included within the meaning of “glycidyl ether.”

The term “hydrogel” means a polymer matrix that swells in water but doesnot dissolve in water.

The term “ether linkage” means an organic functional group composed of acarbon-to-oxygen-to-carbon bond. An example contains a C—O—C linkage.

The term “ester linkage” as used herein for polymerization means anorganic fractional group composed of a O═C—R group, where R is apolymer.

The term “transmural pressure” means the hydrostatic pressure inside thesac minus the hydrostatic pressure outside the sac.

The term “Laplace's laws” refer to two relationships between transmuralpressure, radius of a containment device (e.g., a “bag” or a “sac”) andthe wall stress. For a sphere, the wall tension “T”=Pressure “P” timesRadius “R,” or T=PR. For a cylinder, T=PR/2.

The term “viscosity” refers to a liquid-like property of a materialhaving a relatively high resistance to flow in response to an appliedforce. Viscosity is a measure of the viscous, or liquid-like, nature ofthe material.

The term “Storage Viscosity” (G′/ω) is the elastic modulus G′ divided bythe frequency (ω),

The term “Dynamic Viscosity” (G″/ω) is the loss modulus (G″) divided bythe frequency (ω).

The term “viscoelastic” means a property of polymeric materials thathave both elastic (solid-like) and viscous (liquid-like) properties.

The term “elasticity” means a rheological property defined as thecontribution of the elastic modulus, G′, to the overall stiffness of thematerial. Elasticity includes Percent Elasticity as a specific example.

The term “Percent Elasticity” is defined as to be equal to:100*G′/(G′+G″).

The term “pseudoplastic” means a rheological property of some polymersolutions characterized by a decrease in solution viscosity atincreasing shear rates.

The term “thixotropy” means a rheological property of some polymersolutions characterized by a time-dependent decrease in solutionviscosity at a constant shear.

The term “Fourier Transform Infrared Spectroscopy,” “FTIR Spectroscopy”or “FTIR” means an analytical technique that is used to detect variousorganic functional groups such as esters, ethers, etc. FTIR is based onabsorbance of infrared electromagnetic radiation by molecules (such asfunctional groups).

The term “carboxymethyl anhydroglucose unit” or “CMAG unit” or “CMAG” isan individual repeat unit of a polysaccharide polymer chain.

The term “CMAG/EP ratio” or “CMAG/EP” means the ratio of CMAGequivalents to epoxide equivalents in the cross-linking reaction.

The term “Multi-Arm PEG” refers to PEGs that are formed around a coremolecule permitting multiple PEG molecules to be covalently bonded tothe core. A multi-arm PEG includes a 4-arm PEG, a 6-arm PEG or any PEGhaving multiple PEGs attached to a core molecule.

The term “Multi-Branch PEG” refers to a single PEG polymer havingin-chain epoxide moieties attached thereto. Multi-branched PEGs may becharacterized by having a particular ratio of epoxide:ethylene oxidemoieties. A fully derivatized multi-branch PEG will have anepoxide:ethylene oxide ratio of 2. However, it should be understood thatmulti-branch PEGs may have epoxide:ethylene oxide ratios of less than 2,and that the ratio, on average, need not be integral in a plurality ofPEG molecules.

The term “phosphate buffered saline” or “PBS” means a solution of watercontaining a phosphate buffer.

The term “base catalysis” means a chemical reaction that is speeded upor enhanced in the presence of a base.

The term “acid catalysis” means a chemical reaction that is speeded upor enhanced in the presence of an acid.

The term “ionic cross-linking” or “ionic crosslinking” is a method ofcombining constituents through ionic bonds.

DESCRIPTIONS OF SPECIFIC EMBODIMENTS

The descriptions of specific embodiments is intended to illustrateaspects of this invention, and is not intended to limit the scope ofthis invention. It can be appreciated that other applications of thecompositions described herein can be developed by persons of ordinaryskill in the art without undue experimentation. All of those embodimentsare considered part of this invention.

PEG/CMC Compositions

This invention includes a variety of compositions having CMCs and PEGs,linked with ether or ester bonds. CMCs are polymers composed of sugarresidues linked together, and each of which may have a carboxyl residueattached to the sugar moiety. There are three (3) potential sites forcarboxylation on each sugar residue of CMC. Because a carboxyl residuecan be chemically reactive, those locations on CMC are potential sitesfor derivatization. By controlling the degree of substitution (“DS” or“ds”) of the CMC, the number of active groups on the derivatized CMC canbe controlled.

FIG. 1 depicts CMC and PEG components of this invention. The portion ofCMC shown includes two cellobiose units (each CMAG Unit having a FW of242 Daltons). A PEGDGE molecule is shown with two epoxide units shown(one circled). PEG can be linked to the CMAG Unit either via etherlinkage (bottom left) or an ester linkage (bottom right).

FIG. 2 depicts CMC with PEG cross-linking via ether linkages. Each endof the PEG molecule is shown bonded to a hydroxyl group of adjacent CMCmolecules.

Uses of PEG/CMC Compositions

PEG/CMC compositions of this invention can be used for one or more ofthe following:

(1) as space filling materials, including those suitable forimplantation as load-bearing compositions placed at locations wherecompressive loads may occur, such as in the spinal cord, foraugmentation or replacement of the nucleus pulposus;

(2) as delivery vehicles for controlled release of bioactive substances,such as drugs, growth factors, active peptides, genes, cells, clottingfactors such as thrombin, and antibiotics hormones includingepinephrine, steroids, anti-inflammatory agents and the like, andvasoconstrictors such as norepinephine and the like;

(3) as delivery vehicles for the localized release of bioactivesubstances, such as drugs, growth factors, active peptides, genes,cells, clotting factors such as thrombin, and antibiotics, hormonesincluding epinephrine, steroids, anti-inflammatory agents and the like,and vasoconstrictors such as norepinephine and the like;

(4) as binders for protein coupling and fatty absorption in both tissueengineering and food industries;

(5) lubrication of joints and medical instruments;

(6) tissue coating and tissue protection from fibrosis, neurotoxins,inflammatory mediators, free radicals and other harmful materials;

(7) anti-adhesion compositions; and

(8) as dermal fillers;

Useful properties include, but are not limited to bioadhesion,bioresorbability, antiadhesion, viscosity, and physicalinterpenetration.

Preparation of PEG/CMC Compositions

The PEG/CMC compositions of this invention can be prepared into one ormore of several forms, including gels, membranes, beads, sponges andcoatings.

The technology to link CMC and other CPSs with PEG-epoxides has widepotential for medical devices and includes two types of compositions.One includes methods to decorate CMCs with PEGs without cross-linkingCMC molecules together. Another includes methods to cross-link CMCmolecules together using PEG-epoxides. Reactions of CMCs with aPEG-epoxide is an addition reaction and therefore produces no knowntoxic by-products. Un-reacted PEG-epoxides can be hydrolyzed to PEGdiols, which are known to be biologically compatible.

In contrast, some prior art cross-linking agents can produce undesirableside products. For example carbiodiimide can cross-link polymers, butbecause their reaction is not a simple addition reaction, reactiveby-products are typically produced, which can react with tissues toproduce unwanted side effects. Such side effects can cause serious sideeffects, especially in uses where the polymer composition is intended toremain in place for prolonged periods of time.

Similarly, other prior art cross-linking agents can exhibit unwantedside effects. For example, acrylates such as diacrylate, dimethacrylate,diacrylamide and dimethacrylamide all produce reactive by-products, insome cases, highly toxic by-products. Similarly, hydrazides, tosylates,thiol-containing CPSs (such as thiolated HA) and photoactivatablecross-linkers generally produce toxic by-products, thereby limitingtheir biocompatibility.

The types of CPSs that can be included in this invention is not limitedto CMC. Rather, carboxyethyl cellulose (CEC), hydroxymethyl cellulose,cellulose and other cellulose derivatives.

Compared to prior art CPSs, CMC can be particularly useful in situationswhere it is desired that the residence time of the CPS in tissue beincreased. For example, hyaluronic acid (HA) is a common CPS present inmany tissues. HA has been used for implantable compositions, but HA canbe degraded by tissue enzymes more readily than CMC. CMC is highlybiocompatible, with little, if any, known side effects, and is welltolerated, even after prolonged exposure to tissues.

Space Filling Materials

PEG/CMC compositions of this invention can be particularly useful tofill voids in tissues resulting from disease or injury. For example,removal of a tumor during a surgical procedure can result in a loss oftissue volume. In situations in which organ or tissue function dependson the shape of the organ or tissue, PEG/CMC compositions of thisinvention can be used to fill the void. Similarly, in injuries, such asexcavating injuries in which tissue volume is lost, PEG/CMC compositionsof this invention can be used to fill those voids.

Load-Bearing Materials

Additionally, in situations in which tissue volume is lost throughdegeneration or other causes, PEG/CMC (PEG/CPS) compositions of thisinvention can be used to decrease adverse effects of such tissue loss.In certain aspects, PEG/CMC compositions of this invention can be usedin load bearing capacities. For example, voids in the bone (“bonevoids”) of a vertebral body (e.g., caused by surgical removal of a tumoror degeneration of a vertebral disk nucleus) can result in pain, loss ofsensation and/or loss of motion or function, due in some cases, tocompression of spinal nerves. PEG/CMC compositions can be made withvarying degrees of cross-linking, and in cases in which there is a highdegree of cross-linking, the compositions can support increased loads.To make such compositions, the CMC can be used having a lower molecularweight, so that higher concentrations of CMC may be used resulting inmore ether and carboxyl groups available for cross-linking. Also, usingPEG-epoxides having lower molecular weights, multi-arm PEGs, and/ormulti-branch PEGs can increase the number of covalent cross-links(“cross-link density”) in the matrix. As the cross-link densityincreases, the ability of the matrix to support a load can increase.Thus, in certain aspects, highly-cross-linked PEG/CPS compositions canbe used as nucleus replacements or to fill other bone voids.

In certain of these embodiments, a PEG/CMC (or PEG/CPS) composition canbe placed in a biocompatible sac or bag within the intervertebral space,other bone void where the disk and/or disk nucleus was present, or inother locations where containment of the composition of this inventionis desired. A bag can be made of a silicone-based polymer, such asSilastic™ or another, less deformable material, such as Mylar™ or othersuitable material. It can be desirable to use a bag that has sufficientstrength to resist breakage under the loads expected to be placed on thebag. Additionally, because both the PEG/CPS composition and the walltension (under Laplace's law) can resist a load, these embodiments canbe used in situations in which relatively large loads are to be borne.

Thus, in some embodiments, a biologically compatible bag (which may havea “one-way” valve to permit introduction of material into the bag butwon't permit unwanted loss of material) of an appropriate size to fitwithin the intervertebral space or other location can be inserted in adeflated condition. Such insertion can minimize trauma to the vertebralbodies and the annulus. Once inserted into place, a PEG/CPS cross-linkedcomposition can be introduced into the bag using a needle. Once in thebag, the PEG/CPS composition can polymerize to form a load-bearingstructure, thereby replacing the lost or damaged nucleus. In embodimentsin which there is frank loss of bone (e.g., due to removal of a tumor),a PEG/CPS composition can be formed to the desired shape (as determinedby x-ray, CAT scanning, or other imaging method). The replacementPEG/CPS load bearing compositions generally can be surgically implanted.Alternatively, a biocompatible bag or sac can be inserted in a deflatedcondition as described above for nucleus replacement, and then filledwith a PEG/CPS composition, which is then permitted to polymerize intothe load-bearing material.

In some embodiments, multiple bags or sacs can be used. In particular,in situations in which high loads are to be expected (e.g., the knee,ankles or other lower extremities, or lower back), the use of multiple,small sacs can be employed to take advantage of the fact of thewell-known Laplace relationship between transmural pressure, radius andwall stress of a closed sac. As used herein the term “transmuralpressure” means the hydrostatic pressure inside the sac minus thehydrostatic pressure outside the sac. Thus, for a given transmuralpressure, a sac having a smaller radius will have lower wall stressplaced upon the sac. In contrast, a larger sac will have larger wallstress at the same transmural pressure because of the larger radius.

Based on the Laplace relationships (one for a sphere; T=PR and anotherfor a cylinder; T=PR/2), in certain embodiments of this invention, onecan use a series of small, cylindrical sacs, each inserted into the bonevoid and filled with PEG/CPS composition of this invention. It can bedesirable to align the longitudinal axes of the small sacs in parallelwith the expected load. Thus, during polymerization, the small sacs cansupport the load better than a single sac having larger radius.

Additionally, in situations in which the annulus is damaged, one canintroduce one or more sacs in the intervertebral space, with one nearthe hole in the annulus. Thus, when filled, the sac near the hole in theannulus can effectively seal the hole and minimize further loss ofintervertebral material. In other embodiments, smaller sacs can beinserted and filled (to support the load) and a larger “plug” sac can beintroduced into the intervertebral space. If this plug sac is theninflated with PEG/CPS composition of this invention, the sac caneffectively plug the hole in the annulus and thus prevent the smallersacs from being extruded.

Vehicles for Controlled Delivery of Bioactive Substances

PEG/CMC compositions of this invention can be used to deliver drugs,biologicals, nutrients or other biologically active agents (bioactivesubstances) to an animal. PEG/CMC compositions can be made incorporatinga bioactive substance therein to provide a controlled-releasecomposition. In general, PEG/CMC compositions having more cross-linkstend to retain bioactive substances more than compositions having fewercross-links. Such is the case for bioactive substances that are releasedfrom a PEG/CMC composition by simple diffusion out of the PEG/CMCmatrix. In some cases in which the bioactive substance is large (e.g.,protein), the bioactive substance may be released by a combination ofsimple diffusion out of the PEG/CMC matrix and by release as the PEG/CMCmatrix is degraded in the body.

Regardless of which type of release occurs, it can be appreciated thatthe release of bioactive agents can be controlled as desired by varyingthe composition of the PEG/CMC composition of this invention. It is notintended that the type of bioactive substance be limited. Rather, anybioactive substance whose release is desired to be controlled can beeffectively delivered using PEG/CMC compositions of this invention.

Vehicles for Localized Release of Bioactive Substances

In situations in which local release of a bioactive substance isdesired, a PEG/CMC composition of this invention can be useful. Suchsituations may apply during tissue healing after surgery, wherelocalized trauma to tissues produces localized inflammation. Thus, aPEG/CMC composition of this invention may contain a vasoactive substanceto control bleeding (e.g., a vasoconstrictor, such as norepinephrine) orto promote hemostasis (e.g., a clotting factor). PEG/CMC compositions ofthis invention can also be useful for localized delivery of toxic agentsin chemotherapy. For example, after tumor resection surgery, it may bedesired to administer locally to the site, a PEG/CMC composition havinga chemotherapeutic agent incorporated therein. Such localizedapplication may permit application of higher concentrations ofanti-tumor medications at the site needed, while reducing systemic sideeffects of traditional intravenous injection.

Binders for Protein Coupling

Other uses include more general uses of proteins, fats and otherbiological substances, whether bioactive or not. Thus, proteins can beincorporated into space filling PEG/CMC compositions of this invention.Such proteins may include collagen, gelatin, of other proteins known inthe art.

Lubricants

PEG/CMC compositions of this invention can be effective lubricants formedical instruments. In situations in which a medical instrument isinserted into a body, there is a likelihood of at least some tissuetrauma resulting. Trauma, even slight trauma can cause tissue damage,and can result in unwanted effects. Such effects include bleeding,inflammation, adhesion formation or scarring. By coating a medicalinstrument with a PEG/CMC composition of this invention prior toinsertion into the body, such trauma can be decreased.

Lubrication can be used for both acute and chronic uses. Thus, for anacute procedure, such as urethral catherization, a PEG/CMC compositionof this invention can be used to decrease pain and other discomfort aswell as ease insertion. By decreasing urethral trauma, use of PEG/CMCcompositions of this invention can decrease post-catheterization sideeffects, including decreasing pain on urination, urethral adhesions, anddecreased likelihood of infection.

PEG/CMC compositions of this invention can also be effective tissuelubricants. In situations in which tissues traumatize adjacent tissues(e.g., in joints), compositions of this invention can be used todecrease friction and thereby decrease localized trauma. Thus, byinjecting a PEG/CMC composition into an affected joint, pain can bereduced. By reducing friction and pain, the usual cascade ofinflammation can be inhibited. By inhibiting inflammation, secondaryadverse effects of inflammation can be decreased. Such adverse effectscan be mediated by macrophages, leukocytes, mast cells, eosinophils, andmononuclear cells among others, each of which can produce bioactivesubstances that can make the situation worse. For example, mast cellscontain potent proteases (e.g., mast cell tryptase and mast cellchymotryptase) that can degrade normal tissue proteins. Additionally, aseries of interleukins can be released from neutrophils, macrophages andother inflammatory cells. Interleukins can be potent chemoattractivemolecules and can recruit other inflammatory cells locally to the area,and can thereby continue adverse side effects of tissue trauma. Finally,tissue trauma can activate neuropeptide-containing nerves (e.g.,c-fibers) known to contain substance P, which is a potent stimulus ofpain pathways. In addition to causing pain, substance P is a potentchemoattractant and stimulator of mast cells.

In certain embodiments, PEG/CPS compositions of this invention can beused to lubricate joints, such as those in the spine. In particular,PEG/CPS compositions can be used to lubricate facet joints betweenadjacent lateral spinous processes. Similarly, compositions of thisinvention are useful as lubricants for other joints, including the knee,shoulder, elbows, wrists, ankles and hips.

Tissue Coatings: Antiadhersion Compositions

PEG/CMC compositions of this invention have wide applicability as tissuecoating agents. PEG/CMC compositions can be used as anti-adhesionpreparations. Adhesions are unwanted attachments of a tissue with anadjacent tissue. Adhesions commonly occur after surgery in which tissuesare damaged as a result of the surgery. Thus, PEG/CMC compositions ofthis invention can be effectively used to provide a physical barrierbetween tissues that would otherwise tend to adhere to each other.PEG/CMC compositions may be membranes, gels or sponges. Manyanti-adhesion uses an described in U.S. Pat. Nos. 5,096,997, 6,017,301,6,034,140, 6,133,325, 6,566,345, 6,869,938 and 7,192,984, each patentexpressly incorporated herein fully by reference as if individually soincorporated.

Dermal Fillers

Augmentation of the skin can be an important factor in recovering frominjury or for cosmetic purposes. For example, with normal aging, skinmay become loose or creases can form, such as nasal-labial (nasolabial)folds. In the face, creases or lines may adversely affect a person'sself esteem or even a career. Thus, there has been a need forcompositions and methods that can diminish the appearance of creases orlines.

Further, there are situations in which loss of tissue can leave anindentation in the skin. For example surgical removal of a dermal cyst,lipoatrophy or removal of a solid tumor can result in loss of tissuevolume. In other cases, injuries, such as gunshot wounds, knife wounds,or other excavating injures may leave an indentation in the skin.Regardless of the cause, it can be desirable to provide a dermal fillerthat can increase the volume of tissue to provide a smoother or moreeven appearance.

Several compositions are available for such purposes. Collagen is oftenused as an injectable material for soft tissue augmentation.Additionally, numerous other materials, including proteins, fats,hyaluronic acid (HA), polyalcohols, and other polymers have been used asinjectable dermal fillers. However, non-cross linked, hydrophilicpolymers such as collagen, gelatin and HA have not performed well andmust be covalently cross-linked to remain in place to be effective. Oneexample is ZYDERM®, which is uncrosslinked bovine collagen, was noteffective as a dermal filler unless it was first cross-linked withglutaraldehyde to convert it to ZYPLAST®. Similarly, HA has not beensufficiently effective as a space filling material when injected orimplanted in the body unless it is first cross-linked.

Compositions of CMC and modified CMC have unique properties that allowsuch compositions to be injected into the skin to fill spaces and toprovide support where support is desired. One example for needed supportis dermal augmentation in the face where dermal and subdermal volume islost due to aging. CMC has a unique property of being an elastic gelwith unique physical properties such as dynamic, plastic and zero shearviscosity, tissue adhesiveness, cohesiveness and flow characteristics.In addition, it can achieve these properties without the requirement ofcovalent cross-linking. CMC is particularly unique because chemicalmodifications of CMC expand the number of physical properties that makeit an ideal injectable polymer for human treatment. For example, changein the degree of substitution has a dramatic effect on thixotropy and onviscosity of the gel. Its biocompatability and viscoelastic propertiesmake it uniquely useful for injection into human skin where it becomes aspace filling, biocompatible polymer.

Other polymers tested for their ability to perform as space filling gelsare polysaccharides that have been used for soft tissue filing areinferior to CMC. For example, HA must be cross-linked to cause it tofunction as an elastic gel. Cross-linking limits its ability to beinjected through narrow gauge needles, because the cross-linkingconverts HA into particles. For example, RESTYLANE® is a productconsisting of cross-linked HA in a compatible solution.

Proteins used for dermal augmentation, such as collagen, also must becross-linked to perform well as dermal fillers. For example, ZYPLAST® isa cross-linked bovine collagen dermal filler.

CMC can be a carrier for additional material for additional material forthe skin, including hydrogel polymers such as PEO and emulsions. CMC canbe used to deliver drugs to the skin, such as antioxidants, retinol,vitamins and growth factors. Covalent cross-linking of polymers convertsthem into particles that diminish their ability to deliver additionalpolymers, liposomes, emulsions or other particulates.

Numerous substances have been tested over the years for augmenting softtissue in the dermis in the face to improve cosmesis by fillingdepressions in the skin (Klein and Elson, The History of Substances forSoft Tissue Augmentation, Dermatological Surgery 26:1096-1105, 2000).This is an area that continues to be studied as there has been noclearly superior material or product (Hotta, Dermal Fillers: The NextGeneration, Plastic Surgical Nursing 24(1):14-19, 2004). These fillersare prepared from several polymers including bovine collagen, porcinecollagen, chicken or bacteria fermented HA, gelatin, all of which arecross-linked covalently to reduce their dissolution time orimmunological reactions. Fillers also include autologous human collagen(cross-linked collagen from the patient), human cadaver dermis(cross-linked human collagen). Additional fillers are those that areinsoluble in the dermis, including PMMA beads, dPTFE (expandedpolytetrofluoroethylene), poly lactic acid, recombinant elastin, andthermoplastics that form gels when injected into humans (Klein andElson, The History of Substances for Soft Tissue Augmentation,Dermatological Surgery 26:1096-1105, 2000). More recently, ceramicparticles (U.S. Pat. No. 5,922,025) and also PMMA microspheres (Lemperleet al, Migration Studies and Histology of Injectable Microspheres ofDifferent Sizes in Mice, Plast. Reconstr. Surg 113(5):1380-1390 (2004)have been used for soft tissue augmentation.

Dermal fillers are used to fill scars, depressions and wrinkles. Dermalfiller substances have various inflammatory responses in the dermisincluding phagocytosis to foreign body reactions depending on thematerial (Lemperle et al., Human Histology and Persistence of VariousInjectable Filler Substances for Soft Tissue Augmentation, AestheticPlast. Surg. 27(5):354-366; discussion 367 (2003). One goal of dermalfillers it to temporarily augment the dermis to correct the surfacecontour of the skin without producing an unacceptable inflammatoryreaction, hypersensitivity reaction or foreign body reaction that causespain, redness or excessive scar formation for a period of time.

One of the first materials to be used for dermal augmentation isZYPLAST® derived form bovine collagen. A newer material used for thisapplication is RESTYLANE® derived from bacteria-produced HA. Becausechallenges include both biocompatibility and persistence in the skin,new dermal fillers are compared to one of the existing products such asZYPLAST® or RESTYLANE® (Narins et al., A Randomized, Double-Blind,Multicenter Comparison of the Efficacy and Tolerabiliyt of RestylaneVersus Zyplast for the Correction of Nasolabial Folds, Dermatol. Surg.29:588-595 (2003).

More recently, CMC has been used with polyethylene oxide (PEO) andmultivalent ions to produce ionically linked materials (U.S. Pat. No.7,192,984, incorporated herein fully by reference).

I General Methods

In the following section, manufacture of CMC/PEG compositions arepresented. However, it should be appreciated that CMC need not be theonly CPS used. Rather, any CPS can be used in manufacture of PEG/CPScompositions in similar fashions without departing from the scope ofthis invention.

A. Manufacture of CPS/PEG Compositions

To manufacture CPS/PEG (or CMC/PEG) compositions of this invention,generally the CPS is dissolved in aqueous medium, such as water, saline,phosphate-buffered saline or other suitable medium. For example,dissolving CMC is aqueous media is generally accomplished by adding apre-weighed amount of powdered, dry CMC into a vessel containing themedium with stirring, such as with a vortex mixer until the CMC iscompletely dissolved. In some embodiments, CMC can be present in aconcentration of from about 1% by weight to about 30% by weight. Inother embodiments, CMC can be present in a concentration of about 3% toabout 15% by weight. The molecular weights of CMC can be in the range offrom about 50,000 D to about 1,000,000 D, alternatively from about 90 kDto about 700 kD.

To react PEG with the CMC, a pre-weighed amount of PEG with epoxidemoieties is added to the CMC solution under stirring, until the PEG isdissolved. Then a catalyst is added to initiate the reaction. Generally,the reaction is permitted to go to completion, and the CMC/PEGcomposition is precipitated from solution, dried, and then reconstitutedin solution for analysis and/or use. In some embodiments, the solutioncan be a physiologically compatible solution, with biocompatible pH,ionic strength, and colloid osmotic pressure. It can be appreciated thatsingle chain, multi branched, or multi-arm PEGs having epoxide moietiesthereon can be used.

PEGDGE can be used in a concentration of about 0.01% to about 20%. Themolecular weights of PEG can be in the range of 200 D to about 50,000 D,and in alternative embodiments from about 500 D to about 8000 D.

In certain embodiments, NaCl can be used in a concentration in the rangeof about 0.001% to about 10%, and alternatively from about 0.01 to about5.0%. In certain embodiments, isotonic saline can be used (e.g., about0.9%).

If desired, multivalent ions can be added to the solution to produceionically linked materials. CaCl₂*H₂O can be used in concentrations offrom about 0.001% to about 50% and in alternatives, from about 0.01% toabout 10%. In other embodiments, CMC/PEG compositions can be made inphosphate buffered saline.

Catalysts can be used to initiate cross-linking. For example, aceticacid and citric acid can be used. Citric acid can be used in aconcentration of from about 0.001% to about 50%, or alternatively fromabout 0.01% to about 10%. Acetic acid can be used in a concentration ofabout 0.001% to about 100%, alternatively about 1% to about 20%. Basecatalysis can be accomplished using NaOH in a concentration of about0.001% to about 80%, alternatively about 5% to about 20%. NH₄OH can beused in a concentration of about 0.001% to about 40%, alternatively fromabout 1% to about 20%.

Additionally, pH can be varied to produce compositions that have somehydrogen bonded components. Useful pH ranges can be in the range ofabout 6.0 to about 7.5. Adjusting pH can be accomplished by immersing aCMC/PEG composition in a buffer solution at the appropriate pH.

In other embodiments, CMC/PEG compositions can be used directly aftermixing the CMC and PEG-epoxide together, without subsequentprecipitation and reconstitution. Thus, a CMC/PEGDGE solution can beprepared and drawn into a syringe, the syringe can then be sterilized(e.g., using steam), and the cross-linking reaction can proceed. Whenthe reaction has occurred, the resulting material can be instilled intoa desired location through a small-gauge (e.g., 29 or 30 gauge) needle.In other embodiments, solutions of CMC and PEG can be sterilized beforemixing.

It can be appreciated that in addition to PEGDGE, other PEG-epoxides canbe used, in fashions similar to those used for PEGDGE. Thus, multi-armPEGs, multi-branch PEGs, or PEG-epoxides having different molecularweights can be used without departing from the scope of this invention.

B. Rheological Methods

Once a CMC/PE composition is prepared, its viscoelastic properties canbe readily determined using equipment and methods known in the art.Small deformation oscillation measurements were carried out with aThermo Haake RS300 Rheometer, Newington, N.H., in the cone and plategeometry. All measurements were performed with a 35 mm/1° titanium conesensor at 25° C. The elastic modulus, G′, and loss modulus, G″, wereobtained over a frequency range of 0.628-198 rad/sec. Tan δ wascalculated as G″/G′.

Sodium carboxymethylcellulose (CMC) was obtained from Hercules andpoly(ethylene glycol) diglycidyl ether (PEGDGE) was obtained fromSigma-Aldrich Corporation. According to manufacturer, CMC A had anaverage Mn of ˜700,000 Da and CMC B had a average Mn of ˜200,000 Da.Rheological measurements were performed on gels prepared at 30 mg/mlsolids concentration in BupH Modified Dulbecco's Phosphate BufferedSaline solution (PBS) purchased from Pierce Chemical (catalog No.28374). The solutions were prepared by stirring or CMC into the PBS atroom temperature for at least two hours. The resultant solutions wereclear and colorless with no solids evident and thus were used withoutfiltration.

CMC/PEG compositions can be sterilized using any conventional method,such as steam sterilization, irradiation or filtration.

C. Determination of Compressive Strength of PEG/CPS Compositions

To determine the compressive strength of PEG/CPS compositions, asuitably shaped piece of material (e.g, 1 inch×1 inch×0.25 inches) canbe prepared and placed on a surface such as a table. Once polymerizationhas occurred, a load (e.g., a known weight) can be placed on thecomposition. The weights can be progressively increased until thecomposition fractures. Alternatively, a composition can be placed in avise, with a pressure gauge inserted, and the load increasedprogressively until the composition fractures.

II Preparation of PEG-Derivatized CPS

CPSs can be derivatized with PEGS to form either non-cross linkedmaterials (“decorated CPSs”) or as cross-linked materials of thisinvention. As used herein for PEG, the term “decoration,” “decorating”and like terms refer to covalent attachment of a PEG via one end of thePEG to one site on a CPS molecule. Because only one end of the PEG isattached to the CPS, the other end of the PEG is unbound. Decorated CPSstherefore, are not cross-linked to other CPS molecules nearby or tothemselves via intra-CPS bonds. Thus, the CPS has side chains of PEG“decorating” the CPS molecule. In other embodiments, the CPS/PEGcomposition can be prepared so that intra-chain and/or inter-chaincovalent bonding can occur.

To ensure that only one end of the linker is coupled to the CPS strand,one can use a molar excess (based on the degree of substitution of theCPS) of the CPS. For example, one can use a molar ratio of activecarboxylic acid groups to linker in the range of about 20-about 50 toprovide a high degree of non-cross-linked CPS or CMC. Alternatively, byusing a lower molar ratio, relatively more cross-linking between CPSmolecules can be achieved. In embodiments in which a highly-cross-linkedCPS is desired, one can use a relatively low (e.g., from less than about0.5 to about 20) molar ratio of carboxyl residues to linkers. It can beappreciated that using a molar excess of linker molecules in a solutioncontaining non-constrained CPS molecules (e.g., a relatively dilutesolution of CPS) can promote derivatization of CPS with littlecross-linking. However, in situations in which CPS molecules areconstrained (e.g., high CPS concentrations) or are tightly packedtogether, there may be an increased tendency for cross-links to formbetween different CPS chains. It can be readily appreciated that workersof skill in the art can select a molar ratio of carboxyl residues tolinker molecules to produce a desired degree of cross-linked CPSs.

CPSs decorated with PEGs typically have lower elasticity and higherviscosity than un-decorated CPSs. Thus, CMC decorated with PEG can bemore adherent to tissues, and additionally may be more useful insituations in which un-decorated CMC may have reduced biocompatabilitycompared to PEG-decorated CMC.

A PEG Decorated CPS

In one series of embodiments, the glycyldyl ether moiety of PEG canreact with either of the two types of reactive moieties on a CPX,namely, a hydroxyl or a carboxyl group. If PEG reacts with a hydroxylgroup, the resulting molecule contains an ether linkage. In cases wherethe PEG bonds with a carboxyl group on a CPS, the resulting moleculecontains an ester linkage.

In situations in which there is a molar excess of PEG relative to thenumber of reactive moieties on the CPS, the favored reaction is betweenone PEG and one CPS, without substantial intrachain crosslinking of theCPSs together. Such “PEG decorated CPSs” have particular usefulness forcompositions in which the elasticity is relatively low, in contrast tocrosslinked PEG/CPS compositions.

Decorating CMC with PEG can increase the biocompatability of thecomposition. Decorating CPS with PEG can also decrease the elasticity ofthe composition, because the derivatized material will have fewer freehydroxyls available (in a CPS with high degree of substitution, d.s.).With fewer reactive groups available, the thixotropy of the materialwill be also be decreased.

B. PEG Cross-Linked CPSs

In situations where a more elastic composition is desired, one cancross-link CPSs together using PEGDGE, in which the relative amounts ofCPS and PEGDGE are more equal. In certain embodiments, one can useincreased amounts of CPS relative to PEGDGE, thereby favoring reactionsin which one PEGDGE molecule forms covalent bonds with two CPSmolecules.

It can be appreciated that because of the two types of reactive moietieson a CPS (e.g., a carboxyl group and an ether group), PEG cross-linkedCPSs can have either ester or ether linkages, or an ester link at oneend of the PEG and an ether link at the other end of the PEG.

1. Effect of Acid or Basic Catalysis

We have also found that the reaction conditions can affect the types ofreactions that occur. For example, acid catalyzed addition of PEGDGE toCPS can produce one type of cross-linked composition, whereas basecatalyzed addition of PEGDGE to CPS can produce another type ofcross-linked composition. Each of these two types of reactions producePEG-cross-linked CPSs having desirable, different properties.

2. Multi-Functional PEGs

In addition to PEGDGE (a “bi-functional” PEG), multi-functional PEGs canbe used. Multi-branch PEGs and multi-arm PEGs can be used to makeCMC/PEG compositions having increased cross-linking. For example, use ofthese types of PEGs can permit more rapid polymerization. Thus, suchmaterials can be prepared shortly before implantation, and afterimplantation, in situ cross-linking can occur to produce alonger-lasting, more elastic material. Such compositions can beparticularly desirable in situations where a void is to be filled, orwhere a disk nucleus has become damaged and spinal nerves are impingedupon by vertebral bodies or spinous processes.

It can be appreciated that one can increase the elasticity and/orstiffness of a CMC/PEG composition by using more reactive species. It isknown that increasing the MWs of CMCs (or CPSs) produces solutionshaving higher viscosity, and may thus be more difficult to manipulate.Conversely, decreasing the MW of the CMC can produce aqueous solutionshaving lower viscosities. One can increase the amount of cross-linkingby using CMCs (or CPSs) having lower molecular weights. By using lowerMW CMCs, one can increase the total mass of CMC that can be effectivelydissolved while maintaining a solution viscosity sufficiently low to beeasily manipulated. Thus, with the addition of an appropriate amount ofPEG epoxide sufficient to provide a CMAG/EP ratio needed to produce across-linked composition having a desired elasticity, the reaction ratecan be increased, permitting more easy preparation of materials.

3. In Situ Cross-Linking

Such rapid cross-linking reactions may be particularly desirable for usein situ, in locations where rapid administration is desired. In someembodiments, the components can be sterilized before use, and the finalpreparation can be prepared from sterile solutions. In these situations,a rapidly cross-reacting mixture can be prepared and then introducedinto the site. From the above discussion, there may be multiple ways ofincreasing the rate of cross-linking reactions. Increasing the amount ofPEGDGE or multifunctional PEG, decreasing the MW of the CMC (or CPS),increasing the amount of initiator or catalyst can either alone or incombination, be used to produce compositions that can polymerizesufficiently rapidly to be useful for situations including use inload-bearing joints, excavating injuries, surgical procedures and thelike. One can also use an in situ polymerizing composition to providelong-lasting dermal filling of the more conventional nature, such asfilling nasolabial folds, crow's feet and other dermal lines.

1. Acid Catalysis

In certain embodiments, to produce acid-catalyzed products of CMC andPEG, we used the materials and conditions shown below in Table 1.

TABLE 1 Reaction conditions for acid catalyzed reaction of CMC withPEGDDE [CMC] CMAG EP CMC mg/mL Catalyst mMol mMol CMAG/EP Results 8266630 1% acetic acid 30.99 8.67 3.6 Drastic increase in elasticity 40° C.82666 30 0.01% Citric acid 30.99 2.17 14.3 Elasticity increased 40° C.90252 50 1% acetic acid 51.65 21.67 2.4 Insoluble material too highly40° C. cross-linked 90252 50 1% acetic acid 51.65 10.83 4.8 Drasticincrease in elasticity 60° C. 90252 50 1% acetic acid 51.65 2.17 23.8Elasticity decreased slightly 0° C.

It can be appreciated that the above reaction conditions areillustrative only, and other conditions (CPSs, acids, pH, ratios ofCMAG/EP and the like) can be used to produce variations of thecompositions of this invention that have variations in elasticity. Ingeneral, we found that using stronger acids (e.g., acetic acid) tendedto produce compositions having increased elasticity. We also found thathigher reaction temperatures tended to produce more elastic materials.Moreover, we found that using CMAG/EP ratios of below about 10, tendedto produce compositions having greater elasticity than compositions madewith higher ratios of CMAG/EP. However, using strong acid (e.g., aceticacid) at high temperature (e.g., 40° C.) and a very low ratio of CMAG/EP(e.g., 2.4) produced a composition that was so highly cross-linked thatit was insoluble.

We also found that both reacted materials have slightly lower complexviscosity at low frequencies than the un-derivatized CMC. At highfrequencies, the un-derivatized CMC had a much higher complex viscositythan the derivatized CMC. There are two components of the magnitude ofthe complex viscosity, the storage viscosity (η″=G′/ω)) and the dynamicviscosity (η′=G″/ω). We found that low frequencies, the dynamicviscosity is much higher for the un-derivatized CMC than for thederivatized CMC. This finding indicates that the loss modules G″ ishigher at low frequencies and has a greater contribution to the complexmodulus at low deformation rates.

2. Base Catalysis

In certain embodiments, we react glycidyl ethers with CMC using basecatalysis. In certain embodiments, sodium hydroxide (NaOH) and ammoniumhydroxide (NH₄OH) can be used. We found that basic catalysis using NaOHproduced cross-linked compositions having little ester, and weretherefore predominately ether-linked compositions. These compositionshave lower elasticity than the un-derivatized CMC. Base-catalyzedmaterials have in general, lower elasticity than un-derivatized CMC. Theelasticity of the gel can be altered by adjusting the epoxide tohydroxyl ratio. As the ratio is decreased, elasticity increases.Conversely, as the epoxide content is increased, the elasticity of thecomposition decreases. In certain of these embodiments,highly-cross-linked compositions are not formed and the reactionproduces PEG Decorated CMC.

In contrast, we found that base catalysis using NH₄OH producedcompositions having some ester formation. NH₄OH catalyzed PEGDGEICMCreactions produce compositions having reduced elasticity and are highlyhydrated (swollen) hydrogels.

3. Neutral Catalysis

In situations in which it is undesirable to expose a tissue to eitheracidic or basic compositions, a PEG/CPS polymer can be made usingneutral conditions. Epoxides can react with carboxyl groups or withether groups under neutral conditions, although the rates of thepolymerization reactions are typically shower than those initiated undereither acidic or basic conditions. However, the rate of reaction can beincreased by increasing the number of reactive moieties available forthe reaction. Thus, using CPSs or PEGs having lower average molecularweights, using multi-branch PEGs, multi-arm PEGs or combinations of eachof these components, one can produce polymer compositions with highdegrees of cross-linking, and therefore, higher compressive strength andhigher elasticity compared to compositions made with bi-functional PEGsand high molecular weight components.

C. Sterilization of CMC/PEG Compositions

As noted above, CMC/PEG compositions may be conveniently sterilizedusing heat. In some embodiments, the composition may be heated in anautoclave or other steam producing apparatus. In some cases, it can bedesirable to prepare a CMC/PEG composition and then place it into adelivery device, such as a syringe. CMC/PEG compositions can be madeusing a “3-step” process, in which: (1) a CMC/PEG mixture is obtained,(2) an initiator is added to start the cross-linking reaction and (3)where the cross-linked material is precipitated and reconstituted.Alternatively, a CMC/PEG composition can be made using a “one-step”process, in which the CMC/PEG solution is made with an initiator andthen placed in a delivery device for sterilization. In these situations,heating not only sterilized the composition, but it also increases therate of cross-linking. After sterilization in situ, the CMC/PEGcomposition is ready to use.

III. Uses of CMC/PEG Compositions

CMC/PEG compositions of this invention can be used as space fillingmaterials, delivery vehicles for bioactive substances, load-bearingmaterials, anti-adhesion compositions and/or lubricants for tissues,joints, medical instruments, dermal fillers, anti other medicalapplications.

Space Filling Materials

In other embodiments, space filling materials can be used to providebulk in internal locations. For example, in situations where a void hasbeen created as a result of removal of internal tissue (e.g., removal ofa sebacious cyst, bullet wound, removal of a localized tumor), in anarea not subject to large movements (e.g., the torso), an implant can bemade having a desired shape and having a desired elasticity. Such spacefilling materials, made according to the principles of this invention,can be highly biocompatible, having long residence times in the body.Such materials can be made in any particularly desired shape. Thus, ifthe void is irregular, the surgeon can shape the implant to match thevoid. After making a surgical incision through the skin, the implant isinserted and the wound sutured. Alternatively, the surgeon can inject acomposition comprising a gel having particles of PEG/CMC materialtherein. After introduction into the void, the material can conform tothe shape of the void, thereby providing a uniform appearing structure.

In still further embodiments, space-filling materials can be containedwithin a biocompatible sac. For example, one can insert a PEG/CMCcross-linked composition into the spine to provide support in situationswhere the nucleus of a vertebral disk has become damaged. By encasingPEG/CMC compositions within a sac, the implanted material can resistcompressive forces, and therefore can be used to avoid nerve pinching, acommon cause of pain in subjects with degenerating disks.

In still further embodiments, PEG/CMC materials of this invention can bedried to form membranes. As described in U.S. Pat. No. 5,906,997(incorporated herein fully by reference), CPS/PEO membranes can be madeby preparing a gel, and then drying the gel. Similar membranes can bemade using PEG/CMC compositions of this invention.

Load-Bearing Materials

In additional aspects, PEG/CPS compositions can be used to support loadswithin the skeletal system. For example, the spine is often a locationwhere degeneration, injury or disease can produce loss of structuralsupport. In particular, in conditions in which the disk is damaged,PEG/CPS compositions of this invention can be readily used. Insituations in which the nucleus pulposus is partially or completelylost, compositions of this invention can be used to replace the losttissue. In some of these embodiments, an elastic, relativelynon-compressible composition can be polymerized before insertion intothe affected area. In other situations, one can administer a compositionof this invention prior to its polymerization, so that the compositionpolymerizes in situ. For example, in situations in which there is frankloss of bone, producing an irregularly shaped defect, a mixture ofcomponents of this invention can be injected. After polymerization, thecomposition can fit well into the defect, thereby providing structuralsupport.

In other situations, compositions of this invention can be placed withinone or more bags or sacs. These embodiments can have increasedload-bearing abilities, due to the facts that: (1) a composition can besupported against compression by the bag or sac, and/or (2) thecomposition has its own load-bearing abilities.

FIGS. 15 a and 15 b depict embodiments of this invention in which aPEG/CPS composition is placed within a bag or sac. FIG. 15 a depicts anembodiment 1500 in which a top portion 1504 of the exterior and a sideportion 1508 of the exterior define an enclosed space. In thisembodiment, the PEG/CPS composition 1512 is depicted within the enclosedspace. FIG. 15 b depicts an alternative embodiment 1501 in which theexterior 1504 of the bag or sac has a spherical shape. PEG/CPScomposition 1512 is shown within the space defined by the exterior 1504.

Antiadhesion Materials

In still further embodiments, compositions of this invention can be usedas antiadhesion materials. Methods for using CMC compositions forantiadhesion purposes are described in U.S. Pat. Nos. 5,906,997,6,017,301, 6,034,140, 6,133,325, 6,193,731, 6,869,938, and 7,192,984.Each of the aforementioned patents are expressly incorporated byreference as if separately so incorporated. It can be readilyappreciated that gels, membranes and other forms of the CMC/PEGcompositions of this invention can be used in similar ways.

Drug Delivery Using CMC/PEG Complexes

It can be readily appreciated that any number of drugs, biologicals andother chemical agents can be delivered using the CMC/PEG composites ofthis invention. Certain agents can be advantageously used for localdelivery, providing desired concentration at a desired site, but whiledecreasing undesirable, systemic effects. Such agents include, but arenot limited to therapeutic proteins, such as thrombin to aid inattaining and maintaining hemostasis, growth factors for bone,cartilage, skin and other tissue and cell types. Some of these peptideand protein growth factors include bone morphogenic protein (BMP),epidermal growth factor (EGF), connective tissue growth factor (CTGF),platelet derived growth factor (PDGF), angiotensin and related peptides,and RGD-containing peptides.

Additionally, locally acting drugs include fungicides, histamine,antihistamine, anti-inflammatory drugs (methotrexate), localanesthetics, angiogenesis promoting drugs (e.g., to treat cardiovasculardisease, and anti-angiogenesis factors (e.g., to treat tumors).

DNA-based therapeutics, including antisense DNA, gene therapeutics andRNA-based therapeutics are also suitably delivered using thecompositions of this invention. These agents can be used to eitherinhibit or promote transcription of endogenous genes, or alternatively,can provide exogenous gene products to promote local treatment.

Locally delivered chemotherapeutic agents can also be delivered. Theseinclude, by way of example only, antibiotics to treat microbialconditions, antifungal agents, antiparasitic agents, anti-neoplasticagents including alkylating agents, anti-metabolites and the like.

It can also be appreciated that various hormones and steroids can bedelivered, as can other, systemically acting drugs, which can bedelivered transmucosally or transdermally. These include IgG, clottingfactors and enzymes for treating mucopolysaccharidosis or otherconditions.

Cardiovascular drugs include vasodilators such as β-adrenoreceptoragonists including terbutaline and low-dose epinephrine,α-adrenoreceptor antagonists including norepinephrine, high-doseepinephrine and the like, and vasodilators including nitroprusside andnitroglycerin.

It can be appreciated that the above descriptions are not intended to belimiting to the scope of the invention. Rather, they are intended to berepresentative of the many different embodiments of the invention.

Lubrication of Joints

Certain aspects of this invention include use of PEG/CPS cross-linkedcompositions to provide lubrication for joints and soft tissues. Insituations in which injury to bone, ligaments, tendons, fascia or othersoft tissues has occurred, healing may not produce a smooth-functioningtissue. For example, damage to facet joints in the spine can result inabnormal alignment of vertebrae, which can lead to further damage to thedisk (annulus or nucleus pulposus). Thus, a PEG/CPS composition of thisinvention can be injected between lateral spinous processes of adjacentvertebrae (which normally can slide past one another during normalmovement). After such injection, the lateral spinous processes can beseparated from each other, and the inherent lubrication afforded by thePEG/CPS composition can decrease further irritation.

In a similar fashion, damage to tendons, ligaments and fascias canproduce pain, swelling and decreased function. This, insertion of aPEG/CPS composition of this invention can improve mobility and candecrease the likelihood for further damage to the tissue.

Tissue Protection

PEG/CMC compositions can also be used to protect tissues from damage.For example, such compositions can protect peripheral nerves, tendons,ligaments, other soft tissues, synovial membranes, joints, and canthereby relieve pain.

For example, tendon and ligament injuries heal more slowly than to othertissues, in part because the blood flow to tendons and ligaments isreduced compared to tissues such as muscles, mesenteries, and the like.Furthermore, a tendon stress-related injury often is accompanied by astress injury to an adjacent ligament. Therefore, healing of bothtissues is required for a return to normal function. However, suchrecovery is often slow, and re-injury is common. Further, even whenhealed, tendons and ligaments tend to heal with scar tissue, which isnot smooth. Thus, even after healing, a previously injured tendon orligament may abrade adjacent tissues and cause either re-injury or slowrecovery processes.

In another example, in the spinal cord, damaged spinous processes orvertebral bodies may abrade adjacent tissues. Additionally, loss of avertebral nucleus can lead to compression of vertebral bodies and canresult in impingement of spinal nerves, often leading to pain and/orparalysis.

A further example can involve damage to peripheral nerves, where softtissue injury, trauma or inflammatory reactions can lead to pain or lossof nerve function. Application of a PEG/CMC composition of thisinvention can decrease inflammatory responses, and therefore candecrease secondary damage caused by inflammatory reactions mediated by,for example, macrophages, leukocytes, mast cells or other types ofinflammatory cells.

Additionally, PEG/CMC compositions of this invention can be useful tominimize joint pain. In numerous conditions, including arthritis,traumatic injury, degeneration of cartilage, and ligament damage, ajoint can become painful. A PEG/CMC composition of this invention can beintroduced into an affected joint to provide lubrication and to protectadjacent tissues from damage caused by movement. For example, in theknee, a PEG/CMC composition can be introduced during an arthroscopicprocedure. In situations in which the joint must bear a load (e.g.,knee, hip, ankle, vertebra), a PEG/CMC composition can be made withparticularly high elasticity.

Dermal Filling

Compositions of this invention are particularly well suited as dermalfillers. As noted above, one of the difficulties with prior art dermalfillers is mismatching of the elasticities of the tissue and the dermalfiller. In situations where the tissue is relatively elastic and thedermal filler is relatively inelastic, lumps can appear where the tissuecan stretch, but the dermal filler does not. Conversely, in situationsin which the elasticity of the dermal filler is higher than that of thetissue, incomplete filling of voids can occur.

Thus, by selecting viscoelastic properties of a dermal filler toapproximate or match the elasticity of the tissue, a better void-fillingmaterial can be produced and used, while minimizing adverse effects oftissue-filler mismatching.

Additionally, as an individual ages, the elasticity of the skin tends todecrease. Thus, in subjects with less elastic skin, one might desirablyuse a dermal filler with lower elasticity than one might use in ayounger individual with more elastic skin. Similarly, certain tissuestend to have different elasticities or different mobilities. Forexample, the skin around facial muscles (e.g., nasolabial folds) may besubject to different stresses than other tissues (e.g., the lips). Thus,one can select dermal fillers having different viscoelastic propertiesfor use in the same subject.

Use of such dermal fillers depends upon the specific need. For example,when used to fill small wrinkles, such as nasolabial folds, or “crow'sfeet” around the eyes, dermal fillers in the form of a uniform gel orsmall particles can be desired. An advantage of using a uniform gel isthat these materials can be injected using very small needles, and canproduce a very smooth filling, particularly well suited for smoothingsmall lines. For use in somewhat larger lines (e.g., nasolabial folds),it can be desirable to use compositions comprising a gel havingparticles of PEG/CMC. Such particles can be made according to methodsknown in the art, and can be made to have desired dimensions. For use innasolabial folds, the particles should be sufficiently small to passeasily through a small needle (e.g., a 25 or 30 gauge needle). Theremainder of the composition can be a PEG/CMC gel having relativelylower viscosity. After injection, the particles can further hydrate inthe tissue, thereby forming a more uniform composition.

It can be appreciated that certain embodiments (e.g., “one-step”)embodiments can provide easy to produce, pre-sterilized compositions ina suitable delivery device (e.g., syringe). Pre-made PEG/CMCcompositions, having desirable elasticity, can be injected directly intothe site using a small gauge (e.g. 25, 26, 27, 28, 29 or 30 gauge)needle.

In each of the above situations, PEG/CMC compositions of this inventioncan be beneficial.

EXAMPLES

The following examples are presented to illustrate certain embodimentsof this invention, and are not intended to limit the scope to theembodiments so illustrated. Rather, workers of skill in the art canmodify or adapt the teachings of this invention to make and use othervariations without undue experimentation. All of those embodiments areconsidered to be part of this invention.

Example 1 Crosslinking of CMC A With PEGDGE at a CMAG/Epoxide Ratio of0.8/1 in Dilute NaOH

To a 400 ml polypentene beaker we added distilled, deionized water (DIW)(250 mL) and NaOH (2.5 g; 60.25 mM). After 5 minutes of stirring at 400rpm, CMC A (5.0 g; 40.83 mM OH) was added and stirring continued for 35minutes at 25° C. Subsequently, PEGDGE, (6.884 g; 6.0 mL; 8.7 mM) wasadded neat and stirring was continued at ambient temperature for 90minutes. After that time, we then heated the mixture to about 70° C. for1 hour and about 90° C. for 2 hours. The mixture was then cooled toambient temperature overnight.

The solution was diluted to a volume of about 250 mL with DIW andneutralized with 3.5 ml of glacial acetic acid. At this point the pH was5.3 and 20% NaOH was added to bring the pH to 6.8. The resulting polymerwas precipitated with IPA, collected and then ground in a blender withisopropyl alcohol/methanol (IPA/MeOH) 1:1 in a volume of about 250 mL.The granular solid was collected and washed three times with about 50 mLof acetone then dried in a hood for 20 minutes and the dried in vacuumat a temperature of about 80° C. For rheological testing, the driedcomposition was reconstituted in phosphate-buffered saline (PBS).

Rheological properties of a 30 mg/ml solution in PBS were determinedaccording to methods known in the art. Elasticities of these PEG/CMCcompositions were increased dramatically as shown in FIG. 3 curve A andas described below.

Example 2 Crosslinking of CMC A With PEGDGE at a CMAG/Epoxide Ratio of35.511 in Dilute NaOH

To a 400 ml polypentene beaker, we added DIW (250 mL) and NaOH (10 g;250 mM) with mechanical stirring. After 5 minutes of stirring at 400rpm, we then added CMC A (7.5 g; 67.3 mM OH) and stirring continued for35 minutes at 25° C. Subsequently, we added PEGDGE (0.23 g; neat; 0.2mL; 0.87 mM) and stirring continued at ambient temperature for 90minutes then heated to a temperature of about 70° C. for 3 hours thencooled to ambient temperature overnight.

The solution was then diluted to a volume of about 250 mL with DIW andneutralized with glacial acetic acid to a pH of 6. The resulting polymerwas precipitated with IPA, collected and then ground in a blender withMeOH 250 ml. The granular solid was collected and washed three timeswith about 50 mL of MeOH, then dried in a hood for 20 minutes, and thedried in vacuo at a temperature of about 60° C.

Rheological properties of a 30 mg/ml solution of this material is shownas curve C in FIG. 3. FIG. 3 depicts a graph of G″/G′ ratio (tan δ) vs.frequency for the reaction products of CMC A with PEGDGE in 1% NaOHsolution at CMAG/EP ratios of 0.8, 2.4, 25.6, and CMC with no EP. Theline with filled triangles (▴), A, is the tan δ vs. frequency for thereaction product with the CMAG/EP ratio of 0.8, the line with the filledcircles (), B, is the tan δ vs. frequency for the reaction product withthe CMAG/EP ratio of 2.4, the line with the un-filled squares (□), C, isthe tan δ vs. frequency for the reaction product with the CMAG/EP ratioof 35.6, and the solid line with no symbol (—), D, is the tan δ forun-reacted CMC with no EP. The data clearly indicates that theun-reacted CMC has the lowest tan δ or is the most elastic material. Inthis case, all of the derivatized gels have higher tan δ than un-reactedCMC and as the CMAG/EP ratio is decreased from 25.6 to 2.4 to 0.8 thetan δ is increasing and the material is becoming less elastic. Since theelasticity of the gels decreases upon reaction with PEGDGE, thisindicates that the CMC is decorated and not crosslinked with PEGDGE inthe NaOH catalyzed reaction. By reacting CMC with PEGDGE in base, thetan δ of the CMC/PEG gels can be controlled by adjusting the CMAG/EPratio.

FIG. 4 depicts a graph of G″/G′ (tan δ) at 0.628 rad/sec versus theamount of Epoxide (in mM) of CMC/PEG compositions shown in FIG. 3. Asthe epoxide equivalents increased, and the CMAG/EP ratio decreased, thetan δ increased and the CMC/PEO gel became less elastic. There was anexcellent correlation between the tan δ of the gel and the epoxideequivalents contained in the gel (R²=0.9907).

FIG. 5 depicts FTIR spectra of the PEG/CMC gels from reaction of CMC Awith PEGDGE at different CMAG/EP ratios. Graph A is of a PEG/CMCcomposition where the CMAG/EP ratio was 2.4. Graph B depicts the FTIRspectrum of material having a CMAG/EP ratio of 0.8. Graph C is the FTIRspectrum of material having a CMAG/EP ratio 35.6. Graph D is the FTIRspectrum of un-reacted CMC. For all of the compositions shown, verylittle ester was formed. We conclude from this series of studies thatunder NaOH catalyzed conditions, CMC and PEG form cross-linked polymersdominantly via ether linkages.

Example 3 Crosslinking of CMC A With PEGDGE at a CMAG/Epoxide Ratio of1.78/1 in Dilute NH₄OH

In this example, to a 400 ml polypentene beaker we added DIW (250 mL)and 20% NH₄OH (5 ml) under constant stirring for 5 minutes at 400 rpm.The resulting solution had a pH of 11. To this solution, we added CMC A(7.5 g; 67.3 mM OH) with constant stirring at 700 rpm for 35 minutes ata temperature of 25° C. At this time, PEGDGE (2.3 g; neat 2 mL; 8.7 mM)was added and stirring continued at ambient temperature for 2 hours andthen another 2 ml (8.7 mM) of PEGDGE was added and stirring continuedfor a further 2 hours. The solution was then neutralized with dilute HClto pH 6.8 and the polymer was precipitated with IPA and collected. Thesolid was ground in a blender with MeOH and the granular solid wascollected and wash three times with about 50 mL of MeOH then dried in ahood for 20 minutes and the dried in vacuo at a temperature of about 60°C.

FIG. 6 depicts graphs of tan δ vs. frequency for un-reacted CMC A(filled circles; ; graph A) and a gel made with PEG/CMC produced byreacting CMC A and PEGDGE with a CMAG/EP ratio of 3.57 with basecatalysis in the presence of dilute NH₄OH, filled squares (▪; B). TheNH₄OH catalyzed reaction produced a highly elastic gel that has a muchlower low frequency tan δ than un-reacted CMC A.

FIG. 7 depicts FTIR spectra of a 30 mL solution of this composition inPBS. un-reacted CMC A (graph A), and a PEG/CMC gel formed under basecatalysis using dilute NH₄OH of CMC and PEGDGE (graph B). Graph Bclearly shows the presence of ester formation with a peak at about 1730cm⁻¹ (dashed arrow).

Example 4 Crosslinking of CMC A With PEGDGE at a CMAG/Epoxide Ratio of3.6/1 in Dilute Acetic Acid

To a 400 ml polypentene beaker we added DIW (250 mL) and glacial aceticacid (HOAc) (2.5 ml; 49 mM). After 5 minutes of stirring at 400 rpm, CMCA (7.5 g; 67.3 mM OH) was added, and stirring continued for 55 minutesat ambient temperature. At this time PEGDGE (2.85 g; 2.5 mL; neat; 5.4mM) was added and stirring continued at ambient temperature (about 40°C.) for 5 hours and the stirring stopped and the solution set in at roomtemperature for 70 hours.

The concentrated mass was diluted with 200 mL of DIW and precipitatedwith IPA (300 ml). The entire mass was ground in a blender with 250 mLof MeOH, and the solid was then suction filtered, washed with acetone,dried in the hood, and further dried at a temperature of 60° C. in avacuum oven.

Rheological properties of these materials are shown in FIG. 8 anddescribed below.

Example 5 Crosslinking of CMC A With PEGDGE at a CMAG/Epoxide Ratio of14.3/1 in Dilute Citric Acid

In this series of studies, to a 600 ml polypentene beaker, we added DIW(250 mL) and citric acid (2.5 g) with constant stirring. After 5 minutesof stirring at 400 rpm, CMC A (7.5 g; 31 mM) was added, and stirringcontinued for 55 minutes at ambient temperature. At this time PEGDGE(2.85 g; neat; 0.5 mL; 1.1 mM) was added and stirring was continued at atemperature of 60° C. for 5 hours. After this time, the stirring wasstopped and the solution set at room temperature for 70 hours. Theconcentrated mass was diluted with 200 mL of DIW, neutralized to pH 6.8with dilute NaOH, and polymer was precipitated with IPA (300 ml). Theentire mass was ground in a blender with 250 mL of MeOH, and the solidwas then suction filtered, washed with acetone, dried in the hood, andfurther dried at a temperature of 60° C. in a vacuum oven.

The rheology data of a 30 mg/ml solution in PBS indicates that theelasticity of the cross-linked material has increased, curve C FIG. 8.

FIG. 8 depicts graphs of G″/G′ ratio (tan δ) vs. frequency for reactionproducts of un-reacted CMC A, solid squares (▪; graph A) CMC/PEGDGEreacted in 1% acetic acid with a CMAG/EP ratio of 16, solid circles (;graph B), and CMC/PEGDGE reacted in 0.01% citric acid with a CMAG/EPratio of 14.3, solid triangles (▴; graph C). The acid catalyzedreactions produced gels that were more elastic than the un-reacted CMCat low frequency. In this case, adjusting the CMAG/EP ratio to between3.6 and 14.3 produced gels that had the same low frequency tan δ ofabout 0.3. The acid catalyzed reaction between CMC and PEGDGE appearedto be crosslinking and not decorating the CMAG units. We found that theelasticity of the cross-linked material was increased compared toun-derivatized CMC.

FIG. 9 depicts FTIR spectra of the acid catalyzed reactions of CMC Awith PEGDGE. Graph A depicts the FTIR spectrum of un-reacted CMC A,graph B depicts the FTIR spectrum of CMC A/PEGDGE reacted in 1% aceticacid with a CMAG/EP ratio of 3.6, and graph C depicts the FTIR spectrumof CMC A/PEGDGE reacted in 0.01% citric acid with a CMAG/EP ratio of14.3. These spectra are all very similar and show very little esterpeak. We conclude that the cross-linking under acidic catalysis withacetic acid produces products linked dominantly via ether linkages.

Example 6 Crosslinking of CMC B With PEGDGE at a CMAG/Epoxide Ratio of4.7/1 in Dilute Acetic Acid

In this series of studies, to a 600 ml polypentene beaker, we added DIW(250 mL) and stirred in glacial acetic acid (2.5 ml). After 5 minutes ofstirring at 400 rpm, the pH was 4. Subsequently, CMC B (12.5 g; 51.7 mM)was added and stirring continued at 700 rpm for 35 minutes at atemperature of 25° C. At this time, PEGDGE (2.93 g; neat; 2.5 mL; 5.56mM) was added and stirring continued at ambient temperature for 2 hoursand then the temperature was increased to about 60° C. for 3 hours. Thesolution was then cooled to room temperature and set overnight. Theconcentrated mass was diluted with 200 mL of DIW, neutralized to pH 6.8with dilute NaOH, and precipitated with IPA (300 ml). The entire masswas ground in a blender with 250 mL of MeOH, and the solid suctionfiltered, washed with acetone, dried in the hood, and further dried at atemperature of 60° C. in a vacuum oven.

The elasticity of a 3% solution of this material in PBS is shown in FIG.10. FIG. 10 depicts graphs of tan δ vs. frequency for un-reacted CMC B(solid squares ▪; graph A) and for CMC B reacted with PEGDGE in 1%acetic acid at a CMAG/EP ratio of 5.1/1 (solid circles ; graph B). TheCMC/PEGDGE gel (graph B) is highly elastic (i.e., has a lower tan δ,indicating that CMC has been cross-linked with PEGDGE.

FIG. 11 depicts FTIR spectra of un-reacted CMC B (graph A) and CMC Breacted with PEGDGE in 1% acetic acid at a CMAG/EP ratio of 5.1 to 1(graph B). The cross-linked CMC/PEGDGE material has some ester formationindicated by the arrow at about 1730 cm⁻¹. We conclude from thesestudies that under these conditions, CMC and PEGDGE react with eachother to produced a cross-linked polymer having at least some esterlinkages.

Example 7 CMC/Multi-Branch PEG Compositions

Compositions of CMC with greater cross-linking can be created using PEGsthat are multi-branched. Such PEGs are available from SunBio (Orinda,Calif.), and are depicted in FIG. 12. FIG. 12 depicts a chemicalstructure of a PEG, in which each of the in-chain carbon atoms has anepoxide residue attached thereto. Such a PEG has an epoxide:ethyleneoxide ratio of 2:1. However, one can use PEGS that are not fullysubstituted with epoxide moieties. Rather, one can use PEGs withdifferent ratios of epoxide:ethylene oxide. Thus, altering the number ofepoxide moieties on multi-branch PEGs can be used to provide increasedchoices, along with the use of CMC or other CPSs having differentdegrees of substitution.

To make a cross-linked CMC/multi-branch PEG composition according tothis example, methods as disclosed in Example 1 can be used. A CMC isselected, a multi-branch PEG is selected, a solution of the CMC is madein aqueous solution and the multi-branch PEG is introduced and thereaction is permitted to proceed.

We find that increasing the number of epoxide residues on a multi-branchPEG increases the amount of cross-linking, increases the residence timeand increases the elasticity of the composition.

Example 8 CMC Compositions Containing Multi-Arm PEGs

To provide additional compositions, CMC and multi-arm PEGs can be usedto create cross-linked compositions. A multi-arm PEG is a moleculecontaining more than one PEG molecule, each of which contains at leastone epoxide moiety. Generally, multi-arm PEGS have a core molecule. Inthe case of a 4-arm PEG, the core can be penta-erythritol. For 6-armPEGs, hexose backbones can be used. In both cases, the core moleculeprovides reactive groups that can be covalently bonded to PEG molecules.In the case of a penta-erythritol core, the resulting 4-arm PEG has astructure shown FIG. 13.

To make a CMC/PEG cross-linked composition using a multi-arm PEG, oneuses methods described in Examples 1 or 7. A CMC is selected andprepared in aqueous solution. Then a multi-arm PEG is selected andintroduced into the CMC solution. The reaction is permitted to proceed,and the cross-linked CMC/PEG composition is then isolated.

Example 9 Uses of Multi-Arm PEG and Multi-Branch PEG/CMC Compositions

CMC/PEG compositions described in Examples 7 and/or 8 can be used insituations in which a stronger cross-linking is desired. CMC/multi-armPEG or CMC/multi-branch PEG compositions are used to increase permanenceof the composition for filling excavating voids in internal tissues andthe skin. Such compositions can polymerize at higher rates than PEGDGEbecause of the increased number of reactive epoxide moieties present.

By selecting PEGs having increased number of epoxide moieties and/orlower molecular weights (e.g., lower numbers of ethylene oxidemoieties), the polymerization rates, tissue residence times andelasticities of CMC/PEG compositions can be increased. Similarly,selecting a CMC having a higher degree of substitution can produceCMC/PEG compositions having higher polymerization rates, tissueresidence times and elasticities.

Example 10 Uses as Replacements for Intervertebral Disk Nucleus

In situations in which it is desirable to provide a vertebral disknucleus replacement, a CMC/multi-branch PEG or CMC/multi-arm PEGcomposition can undergo an in situ polymerization reaction. Thus, acomposition having a relatively low viscosity can be implanted into avertebral space. Subsequently, a polymerization reaction can occur,thereby providing a cross-linked, highly elastic composition that can beweight bearing and therefore can reduce symptoms usually associated withextrusion or degeneration of a disk nucleus.

Example 11 Cross-linking of CMC C With PEGDDE at a CMAG/Epoxide Ratio of2.3:1 in Dilute Citric Acid with Steam Sterilization

In this example, we wished to determine if a PEG/CMC composition couldbe made without precipitation and reconstitution before use. Wetherefore carried out a series of studies using CMC/PEGDGE mixturesloaded into syringes without carrying out these steps. We compared theelasticity of such a material with a CMC/PEO composition containingcalcium ions. Such calcium/CMC/PEO compositions are described in U.S.Pat. No. 6,869,938, incorporated herein fully by reference.

We dissolved calcium chloride (0.04 g) and sodium chloride (0.313 g) in50 mL of a solution containing 0.01% citric acid in a 100 mL beaker withmechanical stirring. We then slowly added solid CMC C (1.804 g; 7.45 mM)over about 2 minutes. After 5 minutes of stirring, we then added PEGDGE(0.75 mL; 0.855 g neat; 3.25 mM) and the mixture was stirred for 2 hoursat room temperature. The homogeneous mixture was then loaded into 3 mLpolypropylene syringes and placed into sterilization pouches and steamsterilized in an autoclave for 32 minutes at 122° C. and cooledovernight to room temperature.

The rheology of this CMC/PEGDGE gel was compared to the rheology of aCMC gel without PEGDGE prepared as above is shown in FIG. 14. FIG. 14depicts graphs of G″/G′(tan δ), vs. frequency for two different gels.Graph A (solid squares) depicts results for a gel produced from CMC Creacted with PEGDGE in 0.01% citric acid with CaCl₂ and NaCl and steamsterilization with a CMAG/EP ratio of 2.3. Graph B (solid line) depictsresults for a gel produced from CMC C and non-functional PEO dissolvedin DIW with CaCl₂ and NaCl that was steam sterilized. We conclude thatthe covalently cross-linked CMC/PEGDGE gel has greater elasticity thanthe CMC/PEO ionically cross-linked material, especially at lowfrequencies.

Example 12 Uses as Replacements for Intervertebral Disk Nucleus II

A patient presents with damage or degeneration of a nucleus of anintervertebral disk. The patient is anesthetized and a portion of thespinal column is revealed. To produce a load-bearing structure in theintervertebral space, a biologically compatible spherical or cylindricalsac is inserted in a deflated condition through a small hole in theannulus and into the intervertebral space of a patient in need thereof.Once in place, a PEG/CMC composition of this invention of this inventionis introduced into the sac. Then, the PEG/CMC composition is permittedto polymerize into an elastic, load-bearing structure.

FIGS. 16 a and 16 b depict use of compositions of this invention as apartial or complete replacement for a nucleus pulposus. FIG. 16 adepicts a top view of embodiment 1600 of this invention. A vertebra hasa vertebral body 1604, a dorsal spinous process 1608, lateral spinousprocesses 1612, and a spinal canal 1616 (where the spinal cord islocated). As shown, annulus 1618 surrounds the space where the nucleushas been lost. However, within annulus 1618, a bag 1620 containing aPEG/CPS composition 1624 of this invention has been placed.

FIG. 16 b depicts a lateral cross-sectional view of the embodiment 1600of this invention as shown in FIG. 16 a. Two adjacent vertebrae areshown. Vertebral bodies 1604 are show connected together by annulus1618. Dorsal spinal processes 1608 are depicted for orientation. Thenucleus has been lost, and in its place, bag 1620 is shown filled with aPEG/CPS composition 1624 of this invention.

Example 13 Uses as Replacements for Intervertebral Disk Nucleus III

A patient presents with damage or degeneration of a nucleus of anintervertebral disk. The patient is anesthetized with local or generalanesthetics and a portion of the spinal column is revealed. The locationof the damage is located, and the annulus is evaluated for integrity. Ifthere is no satisfactory hole in the annulus, a small incision in theannulus is made. Multiple PEG/CPS-filled sacs are introduced in eitherinflated or deflated conditions, depending on the size of a hole in anannulus of a patient in need thereof. Where the annulus is intact, asmall hole is made in the annulus and a plurality of cylindrical orspherical sacs is introduced into the intervertebral space. Thelongitudinal axis(es) of a cylindrical sac is aligned parallel to thedirection of the load, and the sacs are then filled with a PEG/CPScomposition of this invention, and the compositions permitted topolymerize.

FIGS. 17 a and 17 b depict an alternative embodiment 1700 of thisinvention. FIG. 17 a depicts a top view of a vertebra having vertebralbody 1604, dorsal spinal process 1608 and lateral spinous processes1612. Spinal canal 1616 is depicted. Annulus 1618 is shown and in thelocation from where the nucleus had been lost has a plurality of smallbags having exteriors 1504 and filled with PEG/CPS compositions 1512 ofthis invention.

FIG. 17 b depicts a lateral cross-sectional view of the embodiment 1700as shown in FIG. 17 a. Two adjacent vertebrae are shown. Vertebralbodies 1604 of each vertebra are shown connected to each other viaannulus 1618, defining a space there in where the nucleus pulposus hasbeen lost. A plurality of bags with exteriors 1504 are shown filled withPEG/CPS composition 1512 of this invention.

Example 14 Uses as Replacements for Intervertebral Disk Nucleus IV

A patient presents with damage or degeneration of a nucleus of anintervertebral disk. The patient is anesthetized with local or generalanesthetics and a portion of the spinal column is revealed. The locationof the damage is located, and the annulus is evaluated for integrity. Ifthere is no satisfactory hole in the annulus, a small incision is madeand one or more cylindrical sacs are introduced into the intervertebralspace, within the annulus. Once so introduced, each sac is then filledwith a PEG/CPS composition of this invention. Then, a larger deflatedsac is introduced into the intervertebral space and is situated near thehole in the annulus. Once positioned, a PEG/CPS composition of thisinvention is introduced into the sac and is permitted to polymerize.This “plug” sac then remains within the annulus and plugs the hole inthe annulus to minimize the extrusion of the smaller sacs through thehole.

FIG. 18 depicts a top view of an alternative embodiment 1800 of thisinvention. A vertebra is shown having vertebral body 1604, dorsal spinalprocess 1608 and lateral spinous processes 1612. Spinal canal 1616 isalso shown. Annulus 1618 is depicted having a defect or hole 1802. Alsoshown is a plurality of bags having exteriors 1504 filled with PEG/CPScomposition 1512 of this invention. Also depicted is PEG/CPS-filled bag1500 shown occluding defect 1802, thereby decreasing the likelihood thatthe bags will be extruded.

It can be appreciated that in the above examples, either a spherical,cylindrical, or other closed shape can be used.

The examples above are provided for purposes of illustrating specificembodiments of this invention. Persons of ordinary skill can readilyproduce other embodiments based on the teachings of this applicationwithout undue experimentation. All such embodiments are included withinthe scope of this invention. All references cited herein areincorporated fully by reference, as is separately so incorporated.

1. A biologically compatible implantable PEG/CMC composition,comprising: a carboxymethyl cellulose (CMC); and a polyethylene glycol(PEG) having at least two epoxide moieties, said PEG linked to said CMCvia an addition reaction of an epoxide moiety to a carboxyl or analcohol moiety of said CMC forming a cross-linked PEG/CMC gel soluble inphysiologically compatible aqueous medium having a carboxymethylanhydroglucose unit (CMAG) to epoxide (EP) ratio of between about 3.6and 14.3, said gel when in solution having a tan δ of <0.5 throughoutthe frequency range of about 0.1 rad/sec to about 100 rad/sec.
 2. Thebiologically compatible implantable PEG/CMC composition of claim 1,where said CMC and a PEG are linked by way of an ether linkage.
 3. Thebiologically compatible implantable PEG/CMC composition of claim 1,where said CMC and a PEG are linked by way of an ester linkage.
 4. Thebiologically compatible implantable PEG/CMC composition of claim 1,wherein the ratio of CMC repeat units (CMAG) to epoxide units is in therange of about 0.5 to about
 30. 5. The biologically compatibleimplantable PEG/CMC composition of claim 1, further comprising anaqueous medium.
 6. The biologically compatible implantable PEG/CMCcomposition of claim 1, where said composition comprises a PEG/CMCcomposition in aqueous suspension.
 7. The biologically compatibleimplantable PEG/CMC composition of claim 1, said PEG/CMC compositionbeing a membrane.
 8. The biologically compatible implantable PEG/CMCcomposition of claim 1, further comprising a drug.
 9. The biologicallycompatible implantable PEG/CMC composition of claim 1, wherein saidcomposition is a bead, solid sphere or sponge.
 10. The biologicallycompatible implantable PEG/CMC composition of claim 1, wherein saidPEG/CMC composition is a PEG/CMC particulate in a PEG/CMC aqueous gel.11. The biologically compatible implantable PEG/CMC composition of claim1, where said PEG is a multiarm PEG or a multibranched PEG.
 12. A methodfor manufacturing a PEG/CMC composition, comprising the steps: selectinga CMC; selecting a polyethylene glycol diglycidyl ether (PEGDDE); mixingsaid CMC and said PEGDDE in aqueous solution; forming a cross-linkedcomposition of said PEG linked to said CMC via an addition reaction ofan epoxide moiety to a carboxyl or an alcohol moiety of said CMC forminga PEG/CMC gel soluble in physiologically compatible aqueous mediumhaving a carboxymethyl anhydroglucose unit (CMAG) to epoxide (EP) ratioof between about 3.6 and 14.3, said gel when in solution having a tan δof <0.5 throughout the frequency range of about 0.1 rad/sec to about 100rad/sec.; and isolating said PEG/CMC composition, where
 13. The methodof claim 12, further comprising adding a catalyst is selected from thegroup consisting of acid catalysts and base catalysts.
 14. The method ofclaim 13, said acidic catalyst being acetic acid or citric acid.
 15. Themethod of claim 13, said base catalyst being sodium hydroxide orammonium hydroxide.
 16. The method of claim 13, said PEG being amultiarm PEG or a multibranched PEG.
 17. The method of claim 12, whereinthe ratio of CMAG to EP is in the range of about 0.5 to about
 30. 18. Abiologically compatible implantable PEG/CMC composition, comprising: acarboxymethyl cellulose (CMC); and a polyethylene glycol (PEG) having atleast one epoxide moiety, said PEG linked to said CMC via an additionreaction of an epoxide moiety to a carboxyl or an alcohol moiety of saidCMC